1. Field of the Invention
This invention relates to the field of biochemistry and to the sub-field of the manufacture of organic solvent- and/or water-insoluble solid supports, and specifically the manufacture of those supports coated with biologically, biochemically or immunochemically active materials (herein referred to henceforth as biomaterials) with retention of one or more of their desired characteristic biological, biochemical or immunochemical functionalities.
2. Description of the Prior Art
Many processes utilize biologically, biochemically, or immunochemically active ingredients attached to insoluble supports. In using such supports, essential steps in many processes may be expedited, may be simplified, may be made less expensive, or improved in other ways. Immobilized biomaterials already appear in literature. Excellent reviews are offered, among others, by R. B. Dunlap, editor, Immobilized Biochemicals and Affinity Chromatography (Plenum Press, New York, N.Y., 1974), and by H. H. Weetall, editor, Immobilized Enzymes, Antigens, Antibodies, and Peptides--Preparation and Characterization (Marcel Dekker, New York, N.Y., 1975). An especially comprehensive review of patented art is found in J. C. Johnson, editor, IMMOBILIZED ENZYMES Preparation and Engineering, Recent Advances, (Noyes Data Corporation, Park Ridge, N.J., 1979).
Messing et al. in U.S. Pat. No. 3,519,538 and Weetall and Yaverbaum in U.S. Pat. No. 4,024,235 each describe a method for bonding biomaterials directly to glass or ceramic supports by incorporating derivatives of silane compounds onto the silicaceous surfaces of a support and chemically coupling biologically or biochemically active molecules through the derivative linkages. The processes described in these patents, however, do not apply to nonsilicaceous materials such as metals, plastics, wood, etc.
Weetall teaches in U.S. Pat. No. 3,652,761 one method whereby organic, biological, or immunological materials can be bonded to glass. It involves the use of an organic silane linker to glass directly, and Weetall demonstrates improved stability of biological materials in this way. Weetall cites as disadvantages of the use of polymers (plastics) a non-specific adsorption and elution, poor flow rates (when used in columns), poor protein coupling efficiency and decreased biological activity. He requires covalent coupling to his glass support. In addition, Weetall shows no applicability to some materials (i.e., metals, nylon, etc.) other than glass.
Glass supports are especially useful in the immunoassay field where antigens and antibodies, respectively, can be bonded to glass media for use in separation steps. Glass is especially desirable as a support medium because of its optical and mechanical properties. In particular, glass tubes (which provide a water-insoluble, water-insuspensible support) bonded to antigens or antibodies are useful because of their relatively low cost and adaptability as cuvettes directly to colorimeters and nephelometers commonly available on the market. A good example of an advanced enzyme immunoassay using such tubes is U.S. Pat. No. 4,016,043 by Schuurs et al. See the teaching of U.S. Pat. Nos. 3,918,817, 3,967,001 and 3,036,492, and the BRICE-PHOENIX Model OM-2000 Light Scattering Photometer (Virtis Co., Gardner, N.Y.) and in particular Ser. No. 932,594, filed Aug. 9, 1978, now U.S. Pat. No. 4,213,764 incorporated herein.
However, glass supports have not always found universal use because of their limited capacity to bond or adsorb biomaterials. The problem is especially acute with respect to certain antigens and antibodies. For example, standard size immunoassay tubes (e.g., 10 mm.times.75 mm, 12 mm.times.75 mm, 13 mm.times.100 mm of glass and coated with thyroxine antibodies take up insufficient antibody to permit an assay in the range desired for most routine analytical testing. There are no commercially available antibody-coated glass tubes on the market now for immunoassays. Commercial clinical immunoassays using other kinds of glass supports are currently sold only by Corning Medical Co., Medfield, Mass., and by Electro-Nucleonics, Inc., of Fairfield, N.J., both of whom purvey assays in which fine glass particles are the supports.
In 1966, K. Catt, H. D. Niall and G. W. Tregar reported in Biochem. J. 100, 31C et seq. (1966) a method for applying serum and immunoglobulins to polymeric supports such that their biochemical functionalities remained essentially intact. Their supports included polystyrene, p-aminobenzylcellulose, and a graft copolymer of polystyrene and polytetrafluoroethylene. Many other workers have since described other polymeric supports with similar properties. See, for example, the work of Ling (U.S. Pat. No. 3,867,517); Tu (U.S. Pat. No. 4,166,844); R. Piasio et al., (U.S. Pat. No. 4,197,287); L. Wide and J. Porath, Biochim. Biophys. Acta 130 at 257 eq seq. (1966); S. A. Tillson et al., in Immunologic Methods in Steroid Determination (F. G. Peron and B. V. Caldwell, editors, Appleton-Century-Crofts, New York, N.Y., 1970); and many papers in Immobilized Biochemicals and Affinity Chromatography, R. B. Dunlap, editor (Plenum Press, New York, N.Y., 1974); and in Immobilized Enzymes, Antigens, Antibodies, and Peptides--Preparation and Characterization, H. H. Weetall, editor (Marcell Dekker, Inc., New York, N.Y. 1975). Again the supports usable with these techniques were limited to polymeric materials.
Catt teaches in U.S. Pat. No. 3,646,346 the use of solid plastic tubes to perform radioimmunoassays. Catt apparently made no attempt to coat glass, and in Catt's teaching, only adsorption of protein was possible.
Bennich et al. in U.S. Pat. No. 3,720,760 taught attaching immunologically active molecules to insoluble polysaccharide (polymeric) beads of the type sold under the trademark SEPHADEX.TM.. Such beads are unstable to swelling upon hydration, their interstices are inherently difficult to wash or rinse, and they have poor mechanical stability (may be crushed, compacted, etc.).
E. A. Fischer has taught by way of U.S. Pat. No. 4,181,636 use of suspensions of polymeric beads for purposes similar to that of Bennich. Neither Bennich nor Fischer described applying the particles to nonsuspendable solid surfaces, which would render them useful without centrifugation. The processes they describe also were limited to the use of polymeric materials.
The substitution of polymeric for nonpolymeric materials will, in many instances, increase the capacity of a solid support to adsorb biomaterials. While, for example, glass will adsorb biomaterials that remain functional, glass supports have not proven as popular in commercial and laboratory applications as have polymeric supports, partly (as indicated above) because their capacity to take up biomaterials is typically less than that of similar surfaces molded from polymeric materials. Many manufacturers offer products similar to glass supports comprising polymers. Mallinckrodt Inc., St. Louis, Mo.; Clinical Assays (Division of Travenol Laboratories, Inc.), Cambridge, Mass.; Squibb, Inc., Princeton, N.J.; Organon Diagnostics, West Orange, N.J.; Abbott Laboratories, Inc., Chicago, Ill.; Cordis Corp., Miami, Fla.; Becton-Dickenson, Inc., Orangeburg, N.Y.; North American Biologicals, Inc., Miami, Fla.; Bio-Rad Laboratories, Richmond, Calif.; RIA Products, Waltham, Mass.; M.A. Bioproducts, Walkersville, Md.; Worthington Diagnostics (Division of Millipore Inc.), Freehold, N.J.; Millipore Inc., Bedford, Mass.; Dade Reagents Inc., Miami, Fla.; Wellcome Reagents, Research Triangle Park, N.C.; Micromedic Systems, Inc.; Horsham, Pa.; Ramco Inc., Dallas, Tex.; Ventrex Inc., Portland, Maine; Litton Bionetics, Kensington, Md.; and New England Immunology Associates, Cambridge, Mass.
Unfortunately, plastic supports have inferior thermal stability vis-a-vis glass--they are amenable to warping by heat and can therefore be deformed. They do not have the desired optical properties of glass. In particular, for fluoresence and enzyme immunoassays, most plastic tubes are very inappropriate because they are opaque or translucent, and/or have optically irregular and inferior surfaces. Because of the increasing price of petrochemicals, their cost is always uncertain. More importantly, supports made of opaque or translucent polymers or polymers with irregular and inferior surfaces cannot be adapted as self-contained cuvettes. In many instances, immunoassays using these supports contain solutions which must be transferred to a glass cuvette. Plastic tubes almost invariably cannot be used with common spectrophotometers.
Hence, an acute need arose in the biochemical field, (particularly the immunoassay art) for a surface to take up biomaterials (e.g., antibodies, antigens, or haptens) which have the adsorptive properties of plastic, but have the same optical and mechanical properties as glass.
It is also known in the art to coat metals with biomaterials. For example, see M. Charles et al. in Immobilized Biochemicals and Affinity Chromatography, R. B. Dunlap, editor (Plenum Press, New York, N.Y., 1974), at 213 et seq. and N. Yamamoto et al., Chemistry Letters (Japan) at 245-246 (1978), who describe methods for applying biomaterials with functional retention of their essential properties to stainless steel, titanium and other dense metals. Methods taught by these authors, however, are by their teachings limited to the materials described, and are not taught to be widely applicable to other support materials like polymers or wood.
S.-P. S. Yen, A. Rembaum and R. S. Molday described in U.S. Pat. No. 4,206,094 a way of preparing extremely small, magentically responsive, polymer-coated particles possessing functional biomaterials. Their process and invention was limited only to metals, metal compounds of electron-dense metals having an atomic number greater than 50 or magnetically attractive metals. The particles were formed in situ by copolymerization of monomers and metal particles. Applicability to macrosupports and to materials other than metals was not shown. Furthermore, the formation of their materials from organic solvents was not demonstrated.
Recently, R. A. Harte described in U.S. Pat. No. 4,133,639 a vessel and handle device prepared from glass or plastic which was useful as a reaction vessel for quantitation of materials in biological solutions. The teaching of Harte requires covalent bonding of active reagents to the walls of the vessel and requires stirring through use of the handle.